The laboratory of Michael Green, where the proposed studies will take place, has recently made a major discovery regarding the development and potential treatment of cancers harboring an activated, oncogenic BRAF mutation (hereafter referred to as BRAF-positive cancers). Using a genome-wide RNA interference screening strategy, they identified 17 genes required for an activated BRAF oncogene (BRAFV600E) to block proliferation of human primary melanocytes. One of the genes they identified encodes a secreted protein, IGFBP7. Several lines of evidence indicate that IGFBP7 is a melanoma tumor suppressor whose loss of expression is required for development of BRAF-positive melanoma. Moreover, addition of exogenous purified, recombinant IGFBP7 selectively kills BRAF-positive human melanoma cell lines and suppresses BRAF-positive melanoma tumor growth in mouse xenografts, suggesting that IGFBP7 may have a role in treating melanoma, as well as other cancers harboring an activating BRAF mutation. The long-term objective of the proposed research is to determine the role of IGFBP7, and the 16 other proteins required for BRAF-mediated apoptosis and senescence, in the development, diagnosis, and treatment of BRAF-positive cancers. The first aim of this project is to continue to study the mechanism by which IGFBP7 kills BRAF-positive human melanoma cells. Towards this goal, experiments are proposed to identify factors that interact with IGFBP7 and are required for IGFBP7 to kill a BRAF-positive melanoma cell. The second specific aim proposes to continue investigating the role of IGFBP7 in treatment of melanoma, both primary and metastatic, as well as other BRAF-positive cancers through the use of a xenograft mouse model. The third aim is to study the molecular basis of melanoma development by analyzing expression of the other 16 proteins identified in the genome-wide RNA interference screen by immunohistochemistry in a panel of BRAF-positive nevi, BRAF-positive melanomas and BRAF-wild type melanomas. The final aim proposes to determine whether IGFBP7, and the other 16 proteins required for BRAF-mediated apoptosis and senescence, might provide useful markers for melanoma diagnosis. PUBLIC HEALTH RELEVANCE: The proposed experiments are designed to investigate the possibility that IGFBP7 may have therapeutic applications in treating melanoma and other malignancies harboring activating BRAF mutations. Moreover, the proposed research seeks to find diagnostic markers that can be used, for example, to distinguish dysplastic nevi (atypical moles) from melanoma, which has important consequences for therapy, prognosis and medical insurability. Thus, the results obtained from these studies will likely have a significant impact on the diagnosis and treatment of melanoma and other BRAF-positive malignancies.